ORISON

ACADEMY

Biology Project
Session : 2021-2022

A Project Report On “ Recombinant

DNA Technology In Todays
Medicine”

Submitted by Shaileshwaran.S
Class – XIIth (Science)
Roll no: (cbse) - ______

Under the Guidance of Mrs.Dhivya

INDEX
 ABSTRACT
 GENETIC ENGINEERING
 STEPS IN SYNTHESIZING A
RECOMBINANT PROTEIN
 GENETIC ENGINEERING HAS PERMITTED
 RECOMBINANT DNA TECHNIQUE
 PLASMID POLYLINKERS AND MARKER
GENES FOR BLUE WHITE SCREENING
 BACTERIAL ARTIFICIAL CHROMOSOMES
AND YEAST ARTIFICIAL CHROMOSOMES
 WHY IS A LENTIVIRUS NECESSARY?
 REFERENCE
 ORISON ACADEMY
Certificate
This is to certify that Shaileshwaran.S student
of class XIIth (Sci.) has successfully prepared
the report on the Project entitled
”Recombinant DNA Technology In Todays
Medicine” under the guidance of Mrs.Dhivya
( Biology).
The report is the result of his efforts & endeavours.
The report is found worthy of acceptance as final
Project report for the subject Biology
of class XIIth (sci.).

Signature of Chemistry Teacher Signature of External

Examiner
--------------------------- --------------------------

Signature of Principal

-----------------------------

Recombinant DNA Technology In

Todays Medicine
Abstract
The goal of this experiment is to study about-Recombinant DNA Technology In
Today’s Medicine.

Genetic Engineering

Genetic Engineering plays a very important role, not only in scientific research,
but also in the diagnosis and treatment of disease. Recombinant DNA is a tool
in understanding the structure, function, and regulation of genes and their
products.
The objectives of Recombinant DNA technology include:
– Identifying genes
– Isolating genes
– Modifying genes
– Re-expressing genes in other hosts or organisms
→ These steps permit scientists and clinicians to:
– Identify new genes and the proteins they encode
– To correct endogenous genetic defects
– To manufacture large quantities of specific gene products such as hormones,
vaccines, and other biological agents of medical interest
→ Genetic engineering produces proteins that offer advantages over proteins
isolated from other biological sources. These advantages include:
– High purity
– High specific activity
– Steady supply
– Batch-to-batch consistency

Steps in Synthesizing a Recombinant Protein

→ Recombinant technology begins with the isolation of a gene of interest. The

gene is then inserted into a vector and cloned. A vector is a piece of DNA that
is capable of independent growth; commonly used vectors are bacterial
plasmids and viral phages. The gene of interest (foreign DNA) is integrated into
the plasmid or phage, and this is referred to as recombinant DNA.
→ Before introducing the vector containing the foreign DNA into host cells to
express the protein, it must be cloned. Cloning is necessary to produce
numerous copies of the DNA since the initial supply is inadequate to insert into
host cells.

→ Once the vector is isolated in large quantities, it can be introduced into the
desired host cells such as mammalian, yeast, or special bacterial cells. The host
cells will then synthesize the foreign protein from the recombinant DNA. When
the cells are grown in vast quantities, the foreign or recombinant protein can
be isolated and purified in large amounts.
→ Recombinant DNA technology is not only an important tool in scientific
research, but has also resulted in enormous progress in the diagnosis and
treatment of certain diseases and genetic disorders in many areas of medicine.

Genetic engineering has permitted

Identification of mutations:

People may be tested for the presence of mutated proteins that may be
involved in the progression of breast cancer, retino-blastoma, and
neurofibromatosis

Diagnosis of affected and carrier states for hereditary diseases:

Tests exist to determine if people are carriers of the cystic fibrosis gene, the
Huntington’s disease gene, the Tay-Sachs disease gene, or the Duchenne
muscular dystrophy gene.
Mapping of human genes on chromosomes:

Scientists are able to link mutations and disease states to specific sites on
chromosomes.
Transferring genes from one organism to another:

People suffering from cystic fibrosis, rheumatoid arthritis, vascular disease, and
certain cancers may now benefit from the progress made in gene therapy.

Isolation and alteration of genes:

Once gene modification becomes successful, alteration of genes to produce a
more functional protein than the endogenous protein may become possible,
opening up the route of gene therapy.

Performing structure and function analyses on proteins:

Researchers may now employ rational drug design to synthesize drug
compounds that will be efficacious and selective in treating disease.

Isolation of large quantities of pure protein:

Insulin, growth hormone, follicle-stimulating hormone, as well as other
proteins, are now available as recombinant products. Physicians will no longer
have to rely on biological products of low purity and specific activity from
inconsistent batch preparations to treat their patients

Recombinant DNA Technique

→ Restriction enzymes used to cut out insulin gene and to cut a bacterial (E.
coli) plasmid at the same “sticky ends”
→ Mutant strains of E. coli used to avoid bacteria attacking “foreign” genes

→ Insert insulin gene next to E. coli

B-galactosidase gene which controls transcription
→ Bacterial cells replicate and make copies of insulin gene
→ Insulin protein is purified (B-galactosidase removed)
→ Chains are mixed and disulfide bridges form
→ Yeast cells provide a sterile growth medium
→ Final product is Humulin - chemically identical to human insulin
Plasmid Polylinkers and Marker Genes for Blue-
White Screening

→ A vector usually contains a sequence (polylinker) which can recognize

several restriction enzymes so that the vector can be used for cloning a variety
of DNA samples.
→ Colonies with recombinant plasmids are white, and colonies with
nonrecombinant plasmids are blue.
→ Example: pUC19
→ Resistant to ampicillin, has (amprgene)
→ Contains portion of the lac operon which codes for beta-galactosidase.
→ X-gal is a substrate of beta-galactosidase and turns blue in the presence of
functional beta-galactosidase is added to the medium.
→ Insertion of foreign DNA into the polylinker disrupts the lac operon, beta-
galactosidase becomes non-functional and the colonies fail to turn blue, but
appear white.
Bacterial Artificial Chromosomes(BACS) And
Yeast Artificial Chromosomes(YACS)

→ BACs can hold up to 300 kbs.

→ The F factor of E.coli is capable of handling large segments of DNA.
→ Recombinant BACs are introduced into E.coli by electroportation( a brief
high-voltage current). Once in the cell, the rBAC replicates like an F factor.
→ Example: pBAC108L
→ Has a set of regulatory genes, OriS, and repE which control F-factor
replication, and parA and parB which limit the number of copies to one or two.
→ A chloramphenicol resistance gene, and a cloning segment.
→ YACs can hold up to 500 kbs.
→ YACs are designed to replicate as plasmids in bacteria when no foreign DNA
is present. Once a fragment is inserted, YACs are transferred to cells, they then
replicate as eukaryotic chromosomes.
→ YACs contain: a yeast centromere, two yeast telomeres, a bacterial origin of
replication, and bacterial selectable markers.
→ YAC plasmid ----> Yeast chromosome
DNA is inserted to a unique restriction site, and cleaves the plasmid with
another restriction endonuclease that removes a fragment of DNA and causes
the YAC to become linear. Once in the cell, the rYAC replicates as a
chromosome, also replicating the foreign DNA.
Why is a Lentivirus necessary?

Lentiviruses can introduce a gene of interest into cells that do not divide –
simple retroviruses cannot.
This ability makes them ideal for a delivery system because most of our cells,
like hemopoietic stem cells, do not divide.

Why use HIV?

A genetically stripped down amalgam of HIV components can be fashioned
with a molecular switch system that turns them off in response to a common
antibiotic.
This type of control allows doctors to control gene expression in people who
are treated with gene therapy - If something goes wrong, the expression can
be turned off.
Adenoviruses are often used as a vector in gene therapy research but they do
not have the capacity to integrate their genome into the hosts genome.
The advantage to using a retrovirus is that you don’t lose the genomic
sequence that is incorporated into the host DNA following cell division.
Reference
→ http://www.rvc.ac.uk/Extranet/DNA_1/DNA_1_intro.htm
→
http://library.thinkquest.org/24355/data/light/details/media/recombinantani
m.html
→ http://www.organoninc.com/products/consumer/
→ NCERT TEXTBOOK
→ GOOGLE
→ PRICIPLES OF GENETICS BY SNUTSON AND BOVERY